Idiopathic Parkinson's disease (PD) afflicts about 1 million Americans, causes widespread suffering in the individuals, and exacts a significant economic toll on society from its production on progressive motor disability. PD symptoms arise from the premature and accelerated death of substantia nigra dopamine neurons and are commonly treated with drugs that replace dopamine synaptic function in the nigrostriatal pathway. However, no available symptomatic drugs have been shown to reduce disease progression by slowing the death of dopamine neurons in humans. Studies performed mainly by the defect in mitochondrial genes coding for complex I of the electron transport chain is responsible for increased oxidative stress, detrimental changes in intracellular calcium signaling, altered mitochondrial replication and movement, and increased susceptibility to cell death mediated by mitochondrial dysfunction. This Project is the third laboratory-based study in a four Project application for a Parkinson's Research Center of Excellence. This Project will explore the molecular mechanisms of cell death produced by PD mitochondria whose genetic functions are isolated and amplified in the cytoplasmic hybrid (cybrid) cell model developed and extensively studied by this group. Results in PD cybrids will be compared to a pharmacological model of chronic complex I impairment produced by rotenone. Four Specific Aims will be examined that test interrelated hypotheses. First, the regulation of mitochondrial membrane potential will be studied with respect to the how cyclical mitochondrial depolarization-repolarization, first described in neural cells by the PI's laboratory, is altered in PD. Second, the effects of over-expressing the anti-apoptotic proteins bcl-2 and bcl-XL on cyclical mitochondrial depolarization and reduced mitochondrial membrane potential in PD will be determined. Third, the molecular mechanisms connecting mitochondrial depolarization to activation of MAPKinase cascade will be defined in control and PD models. Fourth, properties of mitochondrial transition pore (MTP) complexes isolated from PD and control brain tissues will be characterized. These properties included the very recent finding from this group that human brain MTP complexes bind cytochrome c. Depolarization-induced release of cytochrome c and the influence of recombinant bcl proteins and BH region mutants will be studied in MTP-liposomes. This project interacts extensively with the other two laboratory projects and will develop model systems to define mechanisms of action for drugs that might be neuroprotective in PD.